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Pranab K Mukherjee, Nicolas G Bazan; NeuroprotectinD1 (NPD1) mediates Sirtuin signaling in cell survival under-uncompensated oxidative stress in Retinal Pigment Epithelial (RPE) cells. Invest. Ophthalmol. Vis. Sci. 2016;57(12):4387. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Cell survival under uncompensated oxidative stress is important for the understanding of early dysfunctions in retinal degenerations. Sirtuin (SIRT1) is a protein found in retinal pigment epithelial (RPE) cells that promotes cell survival during cellular stress. NPD1, a DHA-derived mediator, protects against neuronal cell death under oxidative stress (OS). This research focuses on the involvement of NPD1-mediated upregulation of Sirtuin in cell survival of RPE cells under stress.
Human RPE (ARPE-19) and primary human RPE cells were used. A programmed cell death (apoptosis) was used to measure the cell death under OS with or without the presence of NPD1. A Western blot (WB) analysis followed for the expression of SIRT1 and 6 in cells under stressed conditions. SIRT1 was silenced by using SIRT1-si RNA in ARPE-19 cells. GFP was used as transfection control. Using these cells, experiments were repeated for apoptosis and WB studies as before.
NPD1 and DHA upregulated Sirtuins (1 and 6) abundance in ARPE-19 cells and in primary human RPE cells undergoing OS. NPD1 displayed a better effect than DHA, however OS or NPD1 alone had little or no effect on SIRT1 and 6 expressions. Moreover, the NPD1-mediated induction of SIRT1 and 6 were time and concentration-dependent. The upregulation of Sirtuins (1 and 6) by NPD1 peaked 6h after initiation of OS, decreased at 12h, and plateaued at 24h in both RPE cells at 100nm of NPD1. Additionally, upregulation of SIRT1 is specific for NPD1, as other structurally-related lipid mediators were ineffective on SIRT1 under similar conditions in RPE cells. Moreover, SIRT1 silencing in ARPE-19 cells under oxidative stress minimalized the expression of SIRT1 in the presence of NPD1 and correlates with programmed cell death under stress.
NPD1 and DHA specifically upregulate SIRT1 and 6 abundance in human RPE cells when confronted with uncompensated OS. As a consequence, remarkable cell survival takes place. The sirtuin silencing ARPE-19 cells devoid of NPD1 sensitivity, allows us to propose a unique DHA/NPD1 survival signaling that plays important role in cell survival under uncompensated oxidative stress.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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